Various vaccines were rapidly developed during the COVID-19 pandemic to prevent and treat infections but global infections continue, and concerns about new mutations and infectious diseases persist. Thus, active research focuses on developing, producing, and supplying vaccines and treatments for various infectious diseases and potential pandemics.Natural killer(NK) cells, as innate immune cells, can recognize and eliminate abnormal cells like virus-infected and cancer cells. Hence, their development as anticancer and antiviral treatments is rapidly advancing. In this study, optimal short-term culture conditions were identified for allogeneic NK cells by simplifying the culture process through the isolation of NK cells(referred to as NKi cells) and eliminating CD3+ cells(referred to as CD3-cells). NK cells demonstrated reduced viral titer in injection of NK cells into SARS-CoV-2 infected ACE-tg mice increased survival.The study's findings could form the basis for an antiviral treatment platform that swiftly responds to new viral disease pandemics.